A clonal tumor cell line (PC12) has been derived from a rat pheochromocytoma. these PC12 cells are an excellent model for studying neuronal differentiation because they display differentiated properties of adrenal chromaffin cells, (PC12-cells) and in addition, acquire neural properties similar to sympathetic neurons after treatment with nerve growth factor (NGF) (PC12+ cells). We have used the PC12 cells and primary sympathetic neurons inculture as an experimental system to study some aspects of differentiation and the biological actions of NGF. We observed that NGF induced the synthesis of a triad of subunits referred to as neurofilament proteins. They are intermediate filaments specific to neurons. Neurofilament triplet proteins in PC12 cells are abnormal when compared to normal neurons in that they are dissociated or incompletely assembled. In addition, PC12 cells are the only clonal cell line in which neurofilament proteins have been found. Recently we have prepared a series of twelve monoclonal antibodies against different antigenic determinants of the neurofilament triplet proteins. We propose to use monoclonal antibodies against neurofilament proteins, together with a combination of biochemical, immunological and immunocytochemical techniques to characterize these intermediate filaments both functionally and structurally following NGF induced differentiation of PC12 cells. Similar studies of the neurofilament proteins in sympathetic neurons will permit comparative observations between data derived from a differentiation tumor cell line which can be induced to differentiate and normal cultured sympathetic neurons. This research proposal will lead to new insights into the development and differentiation of neurons and expand our understanding of neuronal-specific macromolecules which may influence a variety of normal cellular functions carried out by neurons.